Non-RVD mutations that enhance the dynamics of the TAL repeat array along the superhelical axis improve TALEN genome editing efficacy
Autor: | Naoya Tochio, Takehisa Dewa, Takashi Saitoh, Shin-ichi Tate, Masaharu Kondo, Yuichi Togashi, Jun-ichi Uewaki, Takashi Yamamoto, Holger Flechsig, Kohei Umehara, Tetsushi Sakuma |
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Jazyk: | angličtina |
Rok vydání: | 2016 |
Předmět: |
Repetitive Sequences
Amino Acid 0301 basic medicine Computational biology Molecular Dynamics Simulation Biology Article 03 medical and health sciences chemistry.chemical_compound Genome editing Transcription (biology) Transcription Activator-Like Effector Nucleases Gene Editing Genetics Transcription activator-like effector nuclease Nuclease Multidisciplinary urogenital system Superhelix Effector Hydrogen Bonding DNA Dynamic Light Scattering FokI 030104 developmental biology chemistry Mutation Chromatography Gel biology.protein Thermodynamics |
Zdroj: | Scientific Reports |
ISSN: | 2045-2322 |
DOI: | 10.1038/srep37887 |
Popis: | Transcription activator-like effector (TALE) nuclease (TALEN) is widely used as a tool in genome editing. The DNA binding part of TALEN consists of a tandem array of TAL-repeats that form a right-handed superhelix. Each TAL-repeat recognises a specific base by the repeat variable diresidue (RVD) at positions 12 and 13. TALEN comprising the TAL-repeats with periodic mutations to residues at positions 4 and 32 (non-RVD sites) in each repeat (VT-TALE) exhibits increased efficacy in genome editing compared with a counterpart without the mutations (CT-TALE). The molecular basis for the elevated efficacy is unknown. In this report, comparison of the physicochemical properties between CT- and VT-TALEs revealed that VT-TALE has a larger amplitude motion along the superhelical axis (superhelical motion) compared with CT-TALE. The greater superhelical motion in VT-TALE enabled more TAL-repeats to engage in the target sequence recognition compared with CT-TALE. The extended sequence recognition by the TAL-repeats improves site specificity with limiting the spatial distribution of FokI domains to facilitate their dimerization at the desired site. Molecular dynamics simulations revealed that the non-RVD mutations alter inter-repeat hydrogen bonding to amplify the superhelical motion of VT-TALE. The TALEN activity is associated with the inter-repeat hydrogen bonding among the TAL repeats. |
Databáze: | OpenAIRE |
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